TRANSCRIPTION FACTORS REGULATING CHORION GENE PROMOTERS

拟议研究的目标是阐明机制。 确定转录因子的组织和时间特异性 激活。将要使用的模型系统是昆虫绒毛发生，一个过程 涉及严格的开发控制，以表达一些 相关基因，每个基因在Eh上都有不同的调控 转录水平。这些研究将集中在病毒的分子生物学上。 DNA-蛋白质相互作用调节绒毛膜基因的表达 分析这类监管相互作用的保守程度 在物种之间。具体目标将是： 蚕蛾绒毛膜顺式调控基因的定位及功能分析 监管要素。体外识别绒毛膜基因启动子的区域 果蝇卵泡细胞核蛋白与果蝇克隆 转录因子将通过DNase I保护和 甲基化干扰研究。这些地区将成为In的目标 体外诱变及其对组织和组织的影响 将在转基因中分析启动子激活的时间特异性 果蝇。 家蚕绒毛膜基因调控基因的克隆与鉴定 表情。将构建蚕蛾毛囊细胞表达文库 筛选与合成的寡核苷酸结合，代表 卵泡核蛋白的识别位点。此外，基因的cDNA 果蝇与绒毛膜基因转录相关的转录因子 将用于序列同源性筛选。代表CDNA的克隆 假定的调控基因将在结构上进行表征并用于 细菌或组织培养表达中蛋白质的产生 系统。 无细胞条件下转录调控相互作用的重建 系统。来自毛囊的转录活性核提取物 组织培养细胞系将添加果蝇和 蚕蛾可能的调节蛋白及其对体外培养的影响 将分析绒毛膜基因启动子和非绒毛膜基因启动子的转录。 物种间的补充研究将特别揭示 转录调控中的功能保守程度 互动。 上述拟议的研究将提供有关分子机制的信息。 参与一类基因的转录调控，在严格的 发育控制。监管的种间保护 相互作用，如在这个昆虫系统中观察到的，在 哺乳动物系统，尽管仅在少数情况下具有广泛的特点。 关于转录的保存程度的基本信息 物种之间的调控相互作用可能与 人类疾病动物模型的发展。

The goal of the proposed research is the elucidation of mechanisms determining the tissue and temporal specificity of transcriptional activation. The model system to be used is insect choriogenesis, a process involving stringent development controls on the expression of a number of related genes, each of which is differentially regulated at eh transcriptional level. The studies will focus on the molecular biology of DNA-protein interactions regulating the expression of chorion genes and the analysis of the extent of conservation of such regulatory interactions between species. Specific aims will be: Localization and functional analysis of silkmoth chorion gene cis- regulatory elements. Regions on chorion gene promoters recognized in vitro by Drosophila follicular cell nuclear proteins and cloned Drosophila transcription factors will be identified through DNase I protection and methylation interference studies. These regions will be the targets of in vitro mutagenesis and the effect of these mutations on the tissue and temporal specificity of promoter activation will be analyzed in transgenic Drosophila. Cloning and characterization of silkmoth genes regulating chorion gene expression. Silkmoth follicular cell cDNA expression libraries will be screened for binding to synthetic oligonucleotides representing the recognition sites of follicular nuclear proteins. In addition, cDNAs of Drosophila transcription factors associated with chorion gene transcription will be used in sequence homology screening. Clones representing cDNAs of putative regulatory genes will be structurally characterized and used for the production of proteins in bacterial or tissue culture expression systems. Reconstruction of transcriptional control interactions in cell-free systems. Transcriptionally active nuclear extracts derived from follicles and tissue culture cell lines will be supplemented with Drosophila and silkmoth putative regulatory proteins and the effect on the in vitro transcription of chorion and non-chorion gene promoters will be analyzed. The interspecies supplementation studies will be especially revealing on the extent of functional conservation in transcriptional regulatory interactions. The above proposed studies will provide information on molecular mechanisms involved in transcriptional regulation of a class of genes under stringent developmental control. Interspecies conservation of regulatory interactions, such as observed in this insect system, are also evident in mammalian systems, although extensively characterized in only a few cases. Basic information on the extent of conservation of transcriptional regulatory interactions between species could be relevant in the development of animal models for human disease.